The present invention relates to a method for purification of influenza virus, and particularly to such method for obtaining effective vaccines against influenza.
Influenza is an infectious disease which rapidly spreads to lead to nation-wide or even world-wide prevalence. For example, there occured world-wide prevalences in 1918-1921 and in 1957. In the former case, there were reported patients of about 600 millions and deaths of exceeding 20 millions throughout the world, including about 24 millions of patients and 390 thousands of deaths in Japan. Statics figures in the past two decades in Japan shows that 0.3 to 8.5 patients per population of 100 thousands have died due to influenza. Taking into consideration the number of patients died from the complications, the mortality rate with influenza will be estimated to be much higher.
The only possible way to pevent such horrible prevalence of influenza is a vacination, in which there is used a vacine produced by purifying influenza virus. A typical conventional process for purifying influenza virus to produce an inluenza vaccine is conducted as follows: ##STR1##
As shown in the above, the conventional process requires sophisticated steps of purifying influenza virus, such as that contained in the allantoic fluid, by use of density-gradient centrifugation after carrying out refinment steps by low-speed centrifugation and chemical techniques. The refining of influenza virus by subjecting the allantoic fluid directly to the density-gradient centrifugation is not feasible since it will result in damage of the density-gradient liquid layer, Thus, the allantoic fluid containing the virus is pretreated by a chemical refining, generally with barium sulfate, by which the virus is purified by being adsorbed onto the surface of barium sulfate. However, such a prerefining technique with barium sulfate is troublesome in handling a large volume of fluid and entails difficulties in disposing of the barium sulfate after use. Other refining techniques, such as ultrafiltration or high-speed centrifugation, may be considered, but they are still troublesome and not suitable for handling a large amount of allantoic fluid.
In addition, the conventional method as mentioned above is not efficient in extracting immunologically active components of influenza virus. It is generally believed that, among the constituent elements of influenza virus, HA (Hemagglutinin) and NA (Neuraminidase), proteins both known as surface antigens, are the immunologically active components, with HA serving as an infection-preventing antigen and NA assisting in the prevention of infection when a vaccine is prepared from influenza virus. However, the conventional refining process simply removes lipids with ether, and a vaccine produced by such process still contains immunologically inactive or scarcely active proteins, nucleic acids, lipids and other substances, in addition to HA and NA which are generally contained in amounts of about 30% and about 2 to 3% , respectively, of the total proteins of influenza virus. The presence of the unessential substances in the vaccine may cause undesirable side effects when the vaccine is administered. Thus, such vaccine has to be administered in a restricted amount, which limits its immunological effectiveness.